Isoprenoid compounds are the most chemically diverse collection of molecules found in nature. There are currenfiy over 55,000 individual structures, with a large collection of different carbon skeletons and funcfional groups, reported in the literature. All forms of life are able to synthesize isoprenoid compounds cfe novo except for a small group of parasific bacteria with very small genomes that apparenfiy rely on their hosts for essential isoprenoid metabolites. Isoprenoid molecules are built from simpler precursors by prenyl transfer reactions. The prenyl transfer enzymes that mediate these reactions catalyze condensation of electron-rich acceptors (A) with allylic isoprenoid diphosphates. In a typical reacfion, Cl of the allylic substrate is joined to the acceptor with concomitant loss of inorganic pyrophosphate (PP|) and a proton. The acceptors contain nucleophilic moieties - carbon-carbon double bonds, aromatic rings, hydroxyl groups, amino groups, and thiol groups - which are alkylated by the electrophilic allylic diphosphates. Isoprenoid carbon skeletons are constructed from five-carbon 3-methy 1-1-butyl building blocks (the isoprene unit) derived from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). In nature, these units can be joined in one of eight different patterns [1]. Four patterns (1'-2, 1'-4, cr-2-3, and cl'-2-3-2') are constructed from two substrates, IPP and an allylic diphosphate or two allylic diphosphates, by prenyl transfer reactions where the acceptor is a carbon-carbon double bond. The other four skeletons (1'-1, 1'-3, c1'-1-2, and 2-1'-3) result from rearrangements ofthe cl'-2-3 structure. A second diverse set of carbon skeletons is formed by cyclization of linear isoprenoid diphosphates containing two (monoterpene), three (sesquiterpene), and four (diterpene) isoprene units through intramolecular versions of the 1'-2, and c1'-2'-3' condensations [2]. The number of possible carbon skeletons generated by cyclizafion increases as the number of double bonds in the linear polyisoprenoid chain of the substrate increases. The enzymes that catalyze cyclizafion reactions often mediate additional cyclizations and rearrangements to generate a large family of mono-, bi-, and tricyclic structures derived from the primary cyclic structures. The primary cyclic structures for cyclizafion of geranyl diphosphate (GPP) and for farnesyl diphosphate (FPP) at the 6-7 and 10-11 double bonds along with some additional rearranged/bicyclic structures from GPP are shown below. The number of possible cyclic structures formed by the subsequent cyclizations and rearrangements grows substantially for the C15 sesquiterpenes and C20 diterpenes. The condensation reacfions are prenyl transfers where the acceptor is a carbon-carbon double bond. The chemical mechanism of these reactions is a dissociative electrophilic alkylation where the allylic cafion formed from the allylic diphosphate alkylates the double bond [1]. Addifional structures are generated by carbocationic cyclopropylcarbinyl rearrangements of the c1'-2-3 diphosphates [2, 3). Many of cyclic mono-, sesqui-, and diterpenes are formed by intramolecular versions of the prenyltransfer reaction. This group includes structures formed by rearrangements and further cyclizations of the carbocationic intermediates generated by the initial cyclization. Another group of cyclic terpenes is formed by proton-initiated cyclizations and will not be addressed directly in this bridging project [4]. FPP synthase catalyzes the basic chain elongation of DMAPP to FPP (Cio). The enzyme is a homodimer composed of all a-helical subunits [5]. Six highly consen/ed motifs seen in all of GPP, FPP, and geranylgeranyl diphosphate (GGPP) synthases are located on 7 antiparallel alpha helices that form a hydrophobic cavity in the center of the protein with two signature aspartate-rich DDxxD motifs near the opening of the cavity. The aspartates bind and activate the diphosphate group of the allylic substrate through bridging Mg2+ ions [6]. This substructure, called the IS fold, constitutes a superfamily of prenyltransferases that catalyze (1) chain elongafion to form a series of C{10}, C{15}, C{20}, C{25}, C{30}, C{35}, C{40}, C{45}, and C{50} all-trans isoprenoid diphosphates, (2) the c1'-2-3 condensation and subsequent 1'-1 rearrangement of FPP and GGPP to give squalene (sterol biosynthesis) and phytoene (carotenoid biosynthesis), (3) cyclization of GPP, FPP, and GGPP to give cyclic mono-, sesqui-, and diterpenes, and (4) eliminafion or substitution reacfions of DMAPP, GPP, FPP and GGPP to give acyclic terpenoid hydrocarbons and alcohols [1, 4). In addition, there are several descripfions of polyfunctional IS enzymes that give a mixture of products. For example, Artemisia tridentata ssp. spiciformis chrysanthemyl diphosphate (CPP) synthase, which shares 75% sequence idenfity with the A. tndentata FPP synthase (C15 chain elongafion), catalyzes Cio 1'-4 chain elongafion between IPP and DMAPP and c1'-2-3 and 1'-2 condensafion between two molecules of DMAPP [1]. Mulfiple products are also seen for some terpene cyclases [4]. Recenfiy, we showed that only two enzymes in the IS-fold superfamily, CPP synthase and squalene synthase, when placed under forcing conditions, have the capability of forming 7 of the 8 carbons skeletons formed by condensafion or rearrangement [1]. Although enzymes that synthesize the 1'-3, 2-1'-3, c1'-1-2, and c1'-2-3-2'skeletons found in nature have not yet been found, it is likely that they contain the IS fold. Thus, the enzymes that synthesize the majority of the carbon skeletons of isoprenoid molecules have related structures and belong to superfamily defined by the IS fold. Structurally unrelated prenyltransferases include protein prenyltransferases, aromatic prenyltransferases, amino acid prenyltransferases, tRNA prenyltransferases, and glyceryl phosphate prenyltransferases.